In addition to transcriptional control, at least two other levels of genetic regulation must operate in tandemly repeated gene clusters: the conservation of sequence homogeneity and the maintenance of reiteration levels. In fact, very little is known of the mechanisms which govern the stability and transcription of tandem gene families, which are known to encode some of the most basic components for cellular function. As a model system for tandem gene families, the histone genes of Drosophila have been the subject of genetic and molecular experiments in this laboratory. The arrangement and sequence conservation in histone repeats and the reiteration levels and compensatory responses of these genes have been studied. The research described in this application continues to isolate unusual arrangements of histone repeats and to study the adjacent non-histone DNA sequences. The assignment of regulatory importance to such sequences has been complicated by the difficulties in monitoring any particular repeat in the presence of many identical, or nearly identical, histone repeats. The ability to manipulate genes in vitro and to subsequently insert ehm back into the genome by transformation promises a novel approach to the elucidation of the regulatory mechanisms which govern tandemly repeated genes. Experiments are described to use recombinant DNA techniques to create a variety of histone repeats which can be readily distinguished from the typical chromosomal respeats and reintroduce these repeats into the genome of Drosophila by transformation. Transformed flies will be examined for location, transcription, and compensatory responses of the inserted repeats. Repeats which result in transformed flies which exhibit typical transcriptional or compensatory controls will serve as starting points for in vitro mutagenesis. Altered repeats will be used to determine which DNA sequences and arrangements are responsible for regulation of the histone locus.